The success of a transplant of an allograft in a host depends on such factors as the antigens on the transplanted tissue that are recognized by the recipient as foreign and can evoke the rejection response, the cells in the recipient's immune system that mediate rejection, and the reactions that modify either the presentation of the foreign antigen or the cellular response. It is known that a significant component of allograft rejection is due to the presence in donor tissue of non-parenchymal cells (passenger leukocytes).
It is also known that the products of the major histocompatibility complex (MHC) play an important role in mediating an attack by the graft tissue against the recipient. The MHC is generally complex because it includes many different loci, each encoding separate cell-surface antigens, and because the loci have extensive polymorphism. The loci of the MHC fall into one of two classes, Class I or Class II, based on their tissue distribution, the structure of the expressed antigens, and their functions. Class I antigens, present on all nucleated cells, serve as the primary targets for cytotoxic T (CD8.sup.+) lymphocytes. Class II antigens are not distributed in the tissue as widely and serve as primary targets for helper T (CD8.sup.+) lymphocytes.
The polymorphic forms of the individual loci of human leukocyte antigen (HLA), the MHC in humans, have been recognized by antibodies and by various in vitro techniques that measure T-lymphocyte recognition. These responses, mediated by the recipients recognition of polymorphism in the donor, correlate with the strong rejection reactions that take place in vivo. Investigation into the cellular basis of graft rejection, using both in vitro and in vivo studies, has revealed that both CD4.sup.+ and CD8.sup.+ lymphocytes participate in the rejection response.
Attempts to prolong the survival of allografts and xenografts after transplantation, both in experimental models and in medical practice, have centered mainly on the suppression of the immune apparatus of the recipient. This treatment has as its aim preventive immunosuppression and/or treatment of graft rejection.
Examples of agents used for immunosuppression include cytotoxic drugs, antimetabolites, corticosteroids, and antilymphocytic serum. Nonspecific immunosuppressive agents found particularly effective in preventive immunosuppression (azathioprine, bromocryptine, methyl prednisolone, prednisone, and cyclosporin A) have significantly improved the clinical success of transplantation. The nephrotoxicity of cyclosporin A after renal transplantation has been reduced by co-administration of steroids such as prednisolone, or prednisolone in conjunction with azathioprine. In addition, kidneys have been grafted successfully using anti-lymphocyte globulin followed by cyclosporin A. Another protocol is total lymphoid irradiation of the recipient prior to transplantation, followed by minimal immunosuppression after transplantation. Treatment of rejection has involved the use of steroids, 2-amino-6-aryl-5-substituted pyrimidines, heterologous anti-lymphocyte globulin, and monoclonal antibodies to various leukocyte populations.
The principal complication of immunosuppressive drugs is infections. Additionally, systemic immunosuppression is accompanied by undesirable toxic effects, e.g., nephrotoxicity when cyclosporin A is used after renal transplantation, and reduction in the level of the hemopoietic stem cells. Immunosuppressive drugs may also lead to obesity, poor wound healing, steroid hypoglycemia, steroid psychosis, leukopenia, gastrointestinal bleeding, lymphoma and hypertension.
In view of these complications, transplantation immunologists have sought methods for suppressing immune responsiveness in an antigen-specific manner so that only the response to the donor alloantigen would be lost. Such specific immunosuppression generally has been achieved by modifying either the antigenicity of the tissue to be grafted or the specific cells capable of mediating rejection. In certain instances, whether immunity or tolerance will be induced depends on the manner in which the antigen is presented to the immune system.
Mason and Morris ("Effector Mechanisms in Allograft Rejection", Ann. Rev. Immunol. 4:119-45 (1986)) have suggested that a significant component of allograft rejection is the consequence of recipient T lymphocyte recognition of alloantigens expressed by immunostimulatory donor dendritic cells ("DC") present within the grafted tissue. It was logical, therefore, for anti-rejection strategies to focus on the modification and/or elimination of these MHC-bearing "passenger leukocytes" as a more selective and less toxic approach to prevent allograft rejection. It has been hypothesized that such treatment results in the depletion of passenger lymphoid cells and, thus, the absence of a stimulator cell population necessary to tissue immunogenicity.
Using a number of techniques, several attempts to diminish the antigenicity of donor tissues prior to transplantation have been made. The main focus of these attempts has been the total depletion of the donor-derived APCs. The effect of organ culture on the immunogenicity of MHC-incompatible allografts has been studied. Extended time culturing of the donor tissue (Lafferty et al., "Thyroid Allograft Immunogenicity Is Reduced after a Period in Organ Culture", Science, 188:259 (1975)) led to the prolongation of graft acceptance across MHC barriers. (Lafferty et al., Transplantation, 22:138-49 (1976); Bowen et al., Lancet, 2:585-86 (1979). Donor tissue has been treated with growth factor, such as TGF-beta (Czarniecki et al., U.S. Pat. No. 5,135,915 issued 4 Aug. 1992), sometimes in combination with extended culture times (Orton, U.S. Pat. No. 5,192,312 issued 9 Mar. 1993).
Furthermore, donor tissue has been treated with UV light Reemtsma et al., U.S. Pat. No. 4,946,438 issued 7 Aug. 1990; and Lau et al., "Prolongation of Rat Islet Allograft Survival by Direct Ultraviolet Irradiation of the Graft", Science, 223:607 (1984)). It has been suggested that UVB radiation may inhibit LC antigen-presenting cell function by preventing the expression of critical co-stimulatory molecules (Tang et al., J. Immunol., 146: 3347 (1991); Simon et al., "Ultraviolet B Radiation Converts Langerhans Cells from Immunogenic to Tolerogenic Antigen-presenting Cells: Induction of Specific Clonal Anergy in CD4+ T Helper 1 Cells," J. Immunol., 146:485 (1991)). Several authors have suggested that the exposure of LC to UVB or psoralen plus UVA radiation (PUVA) causes a loss of surface markers (including ATPase and class II MHC antigens) without causing overt cytotoxicity (Aberer et al., J. Invest. Dermatol. 76:202 (1981); Hanau et al, J. Invest. Dermatol. 85:135 (1985)). However, Tang and Udey (Tang et al, J. Invest. Dermatol., 99:83 (1992)) showed that the levels of UV radiation that inhibited LC accessory cell function and selectively modulated ICAM-1 expression in short-term cultures were ultimately cytotoxic for LC.
Sometimes UV light has been used in conjunction with microencapsulation (Weber et al., U.S. Pat. No. 5,227,298, issued 13 Jul. 1993). Other workers have used barrier membranes alone, such as the bilayer comprising a first non-cytotoxic layer and a second outer layer of a biocompatible and semipermeable polymeric material taught by Cochrum, U.S. Pat. No. 4,696,286 issued 29 Sep. 1987.
Donor tissues has been treated with a wide variety of substances, such as the topical application of cyclosporin to skin grafts, as disclosed by Hewitt et al., U.S. Pat. No. 4,996,193 issued 26 Feb. 1991, and the perfusion of a donor kidney with lymphocytic chalone, as described by Jones et al., U.S. Pat. No. 4,294,824 issued 13 Oct. 1981. The survival time of skin grafts has been prolonged by treatment in vitro with cortisone, thalidomide, or urethane before implantation into a laboratory animal. The amount of drug locally applied to the skin is usually smaller than the amount required to achieve a similar effect by injecting the drug systemically into the recipient. Donor skin has been treated in vitro with streptokinase/streptodornase, RNA and DNA preparations of the recipient, or solutions of glutaraldehyde, prior to transplantation to reduce the antigenicity of the skin to be grafted.
More sophisticated approaches have involved the treatment of donor tissue with a monoclonal antibody directed against the MHC product along with complement (Faustman et al., "Prolongation of Murine Islet Allograft Survival by Pre-treatment of Islets with Antibody Directly to Ia Determinants", Proc. Natl. Acad. Sci. USA, 78:5156 (1981)) or the treatment of donor tissue with an immunoconjugate of antibody directed against the MHC (Shizuru, et al., "Inhibition of Rat Mixed Lymphocyte Pancreatic Islet Cultures with Anti-Ia Immunotoxin", Transplantation, 42:660 (1986)). Variable results were obtained by these methods.
However, based on the recent observation that microchimerism can exist for many years in the tissues of human solid organ allograft recipients (Starzl et al. "Chimerism and Donor-specific Nonreactivity 27 to 29 Years after Kidney Allotransplantation," Transplantation, 55:1272 (1993)), it has been hypothesized, albeit with much controversy, that microchimerism leads to a state of donor-specific tolerance. Starzl et al., "Liver Transplants Contribute to their Own Success", Nature Med., 2:163 (1996). Since the migratory donor cells required to achieve microchimerism appeared to be the bone marrow-derived dendritic cells (Thomson et al., "Identification of Donor-derived Dendritic Cell Progenitors in Bone Marrow of Spontaneously Tolerant Liver Allograft Recipients", Transplantation, 60:1555 (1995)), it has been recognized that the total depletion of donor-derived DC may not be the best way to achieve the much desired donor-specific tolerance in cell, tissue or organ transplantation. For example, the findings of Rouabhia et al. (Rouabhia et al,. "Cultured Epithelium Allografts: Langerhans Cell and Thy-1.sup.+ Dendritic Epidermal Cell Depletion Effects on Allograft Rejection", Transplantation, 56:259 (1993) suggested that the depletion of Langerhans cells (LC) might not be sufficient to sustain skin and epidermal sheet allograft survival.
The technique employed according the present invention for selectively depleting or attenuating these antigen presenting cells involves contacting donor tissue with a photosensitizer, followed by exposure to light, and then transplantation. Previously, photodynamic methods have-been used primarily for destroying tissues such as tumor tissues, atherosclerotic plaques, surface skin diseases, and unwanted pathogens in blood (Levy et al., U.S. Pat. Nos. 5,283,255 issued 1 Feb. 1994; 4,883,790 issued 28 Nov. 1989; 4,920,143 issued 24 Apr. 1990; 5,095,030 issued 10 Mar. 1992; and 5,171,749 issued 15 Dec. 1992, the disclosures of which are hereby incorporated by reference). See also, Dougherty et al., U.S. Pat. Nos. 4,932,934 issued 12 Jun. 1990; 4,889,129 issued 26 Dec. 1989; 5,028,621 issued 2 Jul. 1991; 4,866,168 issued 12 Sep. 1989; 5,145,863 issued 8 Sep. 1992; and 4,649,151 issued 10 Mar. 1987, which are also hereby incorporated by reference. The capacity of photosensitizers and light to destroy cancerous tissues and unwanted neovasculature constitutes the classical application of photodynamic therapy. Cell death results from either necrotic or apoptotic processes.
For example, U.S. Pat. No. 4,866,168 to Dougherty et al. discloses a composition sold under the trademark "Photofrin II", which is obtained by recovering the high aggregate-molecular weight portion of hematoporphyrin derivative. As another specific example, U.S. Pat. No. 4,883,790 to Levy et al. discloses the use of a group of related compounds designated "monohydrobenzoporphyrins" for analogous purposes.
In addition, the use of many various photosensitizers of similar structure has been described. See, for example, the derivatives of (1-hydroxyethyl)deuteroporphyrin, hydrophobic hematoporphyrin ethers, and compounds prepared from methyl pheophorbide (Pandey et al., U.S. Pat. No. 5,002,962 issued 26 Mar. 1991); pyropheophorbide conjugates (Pandey et al., U.S. Pat. No. 5,314,905); bacteriochlorophyll-a derivatives (Dougherty, U.S. Pat. Nos. 5,171,741 and 5,173,504); monovinyl and divinyl ether-linked dimers (Ward, U.S. Pat. No. 4,961,920); benzoporphyrin derivatives (Allison et al., U.S. Pat. No. 5,214,036); dibenzoporphyrin compounds Dolphin et al., U.S. Pat. Nos. 5,308,608 and 5,149,708); the so-called "green" porphyrins, such as monobenzoporphyrin derivatives (Jamieson et al., U.S. Pat. No. 5,087,636); porphyrin compounds containing exocyclic double bonds (Chang et al., U.S. Pat. No. 5,064,952); and porfimer sodium compositions (Clauss et al., U.S. Pat. No. 5,244,914). The disclosures of all of these patents are hereby incorporated by reference. In general, these drugs are regarded, in a first approximation, as being interchangeable in their utility with respect to photodynamic therapy.
While photodynamic therapy primarily concerns the treatment of tumor cells, additional applications have previously been shown. For example, these photosensitizing drugs can be used in protocols to eliminate atherosclerotic plaques, and in the treatment of blood and of other body fluids to destroy infectious organisms. Further, it has been shown that there is potential to use photodynamic therapy ("PDT") as an immunomodulatory technology for the treatment of a variety of autoimmune conditions. See Richter et al., "Activation of Benzoporphyrin Derivative in the Circulation of Mice without Skin Photosensitivity", Photochem. Photobiol. 59:3, 350-55 (1994); Chowdhary et al., "The Use of Transcutaneous Photodynamic Therapy in the Prevention of Adjuvant Enhanced Arthritis in MRL-1pr Mice", Clin. Immunol. Immunopathol. 72:2, 255-63 (1994); Hunt et al., "Transcutaneous Photodynamnic Therapy Delays the Onset of Paralysis in a Murine Multiple Sclerosis Model", Proc. Soc. Photo-Optical Instr. Eng., 2371:451-55 (1994); and Obochi et al., "Targeting Activated Lymphocytes with Photodynamic Therapy: Susceptibility of Mitogen-stimulated Splenic Lymphocytes to Benzoporphyrin Derivative (BPD) Photosensitization", Photochem. Photobiol. 62:1, 169-75 (1995). It has been shown that exposure of freshly isolated murine Langerhans cells ("LC") to psoralen plus ultraviolet light A (UVA) radiation in vitro inhibited the accessory cell function of LC by decreasing their expression of the intracellular adhesion molecule-1 (ICAM-1). Tang et al., "Effects of Ultraviolet Radiation on Murine Epidermal Langerhans Cells: Doses of Ultraviolet Radiation that Modulate ICAM-1 (CD54) Expression and Inhibit Langerhans Cell Function Cause Delayed Cytotoxicity in vitro", J. Invest. Dermatol., 99:83-89 (1992).
While some workers in this field have shown that transdermal PDT profoundly suppressed contact hypersensitivity (CHS) and can enhance the length of skin allograft acceptance (Simkin et al., "Effect of Photodynamic Therapy Using Benzoporphyrin Derivative on Cutaneous immune Response", Proc. Soc. Photo-Optical Instr. Eng., 2392:23-32 (1995)), the specific cell targets have not been identified as yet for this kind of treatment. Possible candidates include activated lymphocytes in the circulation, activated macrophages or dendritic cells in the circulation, keratinocytes, and Langerhans cells ("LC") in the skin. It has now been found that the imnmunomodulatory effects of "low-dose PDT" of tissue grafts associated with extended engraftment may depend on a selective effect upon epidermal Langerhans cells ("LC") and may not require complete cell depletion, i.e., the eradication of passenger leukocytes, to permit acceptance of the allograft.
It is a particular advantage of the present invention that, unlike therapeutic regimens involving the administration of a photosensitizing drug to an organism, donor tissue may be most appropriately treated in vitro prior to an actual transplant procedure. In this manner, problems associated with ensuring proper levels of light exposure of, e.g., a conjugate associated with target cells within an organism, are substantially obviated.
Further, the method of the invention results in grafts that are immunologically stable in suitable hosts, biologically functional, and capable of being stored prior to transplantation. Thus, this invention enables the establishment of a bank of photodynamically treated grafts that can be used for short-term storage.